Faulstich H, Tews P, Heintz D
Max-Planck-Institut für Medizinische Forschung, Heidelberg, Germany.
Anal Biochem. 1993 Feb 1;208(2):357-62. doi: 10.1006/abio.1993.1061.
n-Octyl-5-dithio-2-nitrobenzoic acid (ODNB), the reaction product of n-octane-1-thiol and Ellman's reagent, can be isolated in crystalline form. Like Ellman's reagent, ODNB can be used for the titration of SH groups in proteins, but with considerable advantages: Due to the absence of one negative charge and the presence of a lipophilic hydrocarbon chain, titrations of protein thiols are significantly faster. Normally, endpoints are reached after 5-30 min, while the corresponding reaction with Ellman's reagent may take 1-2 h. Another advantage of ODNB is that it finds access to thiol groups which normally remain undetected. With myosin S1, for example, ODNB reacts with a thiol group which is not reactive with Ellman's reagent. Although ODNB has detergent properties, reaction of this thiol group as a consequence of denaturation seems unlikely, because ODNB was used below its critical micelle concentration and all other thiols of myosin S1 remained buried. The n-octylthiol derivatives of proteins formed by reaction with ODNB can be used for transient protection of thiols, for example, during purification procedures, or for a reversible blocking of SH groups involved in biochemical reactions.
正辛基-5-二硫代-2-硝基苯甲酸（ODNB）是正辛烷-1-硫醇与埃尔曼试剂的反应产物，可分离得到结晶形式。与埃尔曼试剂一样，ODNB可用于滴定蛋白质中的巯基，但具有相当大的优势：由于缺少一个负电荷且存在亲脂性烃链，蛋白质硫醇的滴定速度明显更快。通常，5-30分钟后即可达到终点，而与埃尔曼试剂的相应反应可能需要1-2小时。ODNB的另一个优点是它能够与通常未被检测到的巯基发生反应。例如，对于肌球蛋白S1，ODNB会与一个对埃尔曼试剂无反应性的巯基发生反应。尽管ODNB具有去污剂性质，但由于该巯基的反应是由变性引起的这一可能性似乎不大，因为ODNB是在其临界胶束浓度以下使用的，且肌球蛋白S1的所有其他巯基仍被掩埋。通过与ODNB反应形成的蛋白质正辛基硫醇衍生物可用于巯基的瞬时保护，例如在纯化过程中，或用于对参与生化反应的巯基进行可逆阻断。
